huvec change medium?? - (Jul/30/2009 )

Hi people...

my friend gave me a vial of PCS-100-010 HUVEC primary cells. Can I use F12K or any other medium with FBS to replace the low-FBS medium that you are supposed to use with this line? I tried using my usual complete medium 10% FBS and the cells did not attach... what should I do?

also, in HUVEC-CS (ATCC), wat does the CS stand for?

thanks...

Chris

my friend gave me a vial of PCS-100-010 HUVEC primary cells. Can I use F12K or any other medium with FBS to replace the low-FBS medium that you are supposed to use with this line? I tried using my usual complete medium 10% FBS and the cells did not attach... what should I do?

I haven't worked with these, but I have worked with HUVECs from commercial sources as well as some just 2 -3 passages from harvest. We always have to coat our plasticware with gelatin, fibronectin, collagen, etc. Gelatin is cheaper so we usually start with it for any primary cells we are culturing. This might help with your attachment problem.

It's been a while since I have cultured them, but I believe we also had to supplement the medium with growth factors. If none of this works, I suggest you ask the person that gave them to you for their protocol on culturing this particular line of HUVECs.



also, in HUVEC-CS (ATCC), what does the CS stand for?

Check out the paper below (sited in the ATCC product lit). It gives information on the derivation of this line.
Gifford SM, et al. Functional characterization of HUVEC-CS: Ca2+ signaling, ERK 1/2 activation, mitogenesis and vasodilator production. J. Endocrinol. 182: 485-499, 2004. PubMed: 15350190

HUVECs did not attach that mean there is no survival cells?

Try
M199+20%FBS+10%(or more)EGM-2 (containing growth factor)

mydove on Jul 31 2009, 06:48 PM said:

hi there,
i've already been culturing my CS in gelatin -coated flasks using F12K medium and ECGS and FBS. My CS cells grw fine. But the PCS cells are primary and supposed to grow in M200 medium with low FBS. Do you think it would be a problem to grow these low-serum cells in the same medium as my previous cells? Or just lower the serum %? Or maybe storage was not good and that's why teh cells did not attach.....

Chris

Perhaps you are right and the freezing or storage killed the cells. Have you tried a viability assay on any of the floating ones or do you know that they are dead? How many vials do you have? You might just try putting them in your other medium or if there are enough cells in one vial, split it into two dishes with either the medium you use for the other ones or the recommended medium.

By the way, we use the same medium as mydove, but we use 200 ul/50 ml Bovine Brain Extract (Clonetics CC-4092) instead of the ECM-2. However, it has been a couple of years since I cultured them so I think Clonetics in now a part of Lonza (CC-4098).

HUVECs require a very specific medium. The ones from ATCC only needs 2% FBS but you need to supplement the medium with an endothelial growth factor kit or they will not be happy. I get what they recommend at ATCC and they grow fine. You may want to check out PCS-100-030 and PCS-100-040/PCS-100-041.